Use of the Ussing chamber technique to study nutrient transport by epithelial tissues

10.2741/4178 ◽  
2013 ◽  
Vol 18 (4) ◽  
pp. 1266 ◽  
Author(s):  
yulong yin
Keyword(s):  
Ussing Chamber ◽  
Nutrient Transport ◽  
Epithelial Tissues ◽  
Chamber Technique

scholarly journals Changes in porcine nutrient transport physiology in response to Ascaris suum infection

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Sarina Koehler ◽  
Andrea Springer ◽  
Nicole Issel ◽  
Stefanie Klinger ◽  
Christina Strube ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Ascaris Suum ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Nutrient Transport ◽  
Transport Processes ◽  
Infected Group ◽  
Peptide Transporter ◽  
Control Group ◽  
Expression Levels

Abstract Background The roundworm Ascaris suum is one of the parasites with the greatest economic impact on pig farming. In this context, lower weight gain is hypothesized to be due to decreased nutrient absorption. This study aims at characterizing the effects of A. suum infection on intestinal nutrient transport processes and potential molecular mechanisms. Methods Three groups of six piglets each were infected orally (10,000 embryonated A. suum eggs) in a single dose (“single infection”). Another three groups were infected orally (1000 embryonated eggs) for 10 consecutive days (“trickle infection”). Animals were necropsied 21, 35 and 49 days post-infection (dpi). Three groups served as respective controls. The Ussing chamber technique was applied for the functional characterization of small intestinal tissues [short-circuit currents (Isc) as induced by glucose, alanine and peptides; 3H-glucose net flux rates; tissue conductance (Gt)]. Transcription and expression levels of relevant cytokines and nutrient transporters were evaluated (qPCR/western blot). Results Peptide- and alanine-induced changes in Isc were significantly decreased in the jejunum and ileum of the trickle-infected group at 49 dpi and in the ileum of the single-infected group at 49 dpi. No significant differences regarding glucose transport were observed between the Ascaris-infected groups and the control group in Ussing chamber experiments. Transcription levels of the glucose and peptide transporters as well as of selected transcription factors (transcription of signal transducer and activator of transcription 6 [STAT6] and hypoxia-inducible factor 1-alpha [Hif-1α]) were significantly increased in response to both infection types after some periods. The transcription of interleukins 4 and 13 varied between decrease and increase regarding the respective time points, as did the protein expression of glucose transporters. The expression of the peptide transporter PepT1 was significantly decreased in the ileal single-infected group at 35 dpi. Hif-1α was significantly increased in the ileal tissue from the single-infected group at 21 dpi and in the trickle-infected group at 35 dpi. The expression levels of Na+/K+-ATPase and ASCT1 remained unaffected. Conclusions In contrast to the current hypothesis, these results indicate that the nutrient deprivation induced by A. suum cannot be explained by transcriptional or expression changes alone and requires further studies. Graphical abstract

Permeability of human jejunal segments to gonyautoxins measured by the Ussing chamber technique

Toxicon ◽  
2004 ◽  
Vol 44 (5) ◽  
pp. 521-528 ◽  
Author(s):  
Pamela Mardones ◽  
Darío Andrinolo ◽  
Attila Csendes ◽  
Néstor Lagos
Keyword(s):  
Ussing Chamber ◽  
Chamber Technique

Transport of selenium species in Caco-2 cells: Analytical approach employing the Ussing chamber technique and HPLC-ICP-MS

2013 ◽  
Vol 110 ◽  
pp. 8-14 ◽  
Author(s):  
Denis Pick ◽  
Christian Degen ◽  
Matthias Leiterer ◽  
Gerhard Jahreis ◽  
Jürgen W. Einax
Keyword(s):  
Analytical Approach ◽  
Ussing Chamber ◽  
Selenium Species ◽  
Icp Ms ◽  
Chamber Technique

Abstract MP10: Furin-Mediated Modification Is Required For Epithelial Sodium Channel-Activating Activity Of Soluble (Pro)Renin Receptor In Cultured Collecting Duct Cells

Hypertension ◽  
2021 ◽  
Vol 78 (Suppl_1) ◽  
Author(s):  
Huaqing Zheng ◽  
Changjiang zou ◽  
Tianxin Yang
Keyword(s):  
Cleavage Site ◽  
Collecting Duct ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Separate Experiment ◽  
Pcr Analysis ◽  
Furin Cleavage ◽  
Empty Vector ◽  
Furin Cleavage Site ◽  
Chamber Technique

(Pro)renin receptor (PRR) contains overlapping cleavage site for site-1 protease (S1P) and furin for generation of soluble PRR (sPRR). Although S1P-mediated cleavage mediates the release of sPRR, the functional implication of furin-mediated cleavage is unclear. Here we tested whether furin-mediated cleavage was required for the activity of sPRR in activating ENaC in cultured M1 cells. M1 cells were transfected with pcDNA3.4 containing full-length PRR with (Furin Mut) or without (WT) mutagenesis of furin cleavage site; empty vector (EM) served a control. As compared with EM, overexpression of WT and Furin Mut vectors induced a more than 16-fold comparable increase in the release of sPRR. Amiloride-sensitive short circuit current as assessed by Ussing chamber technique was elevated by overexpression of WT PRR which was reduced by 37% by Furin Mut (ENaC activity: 1.00 + 0.06 μA/cm 2 in EM, 1.67 + 0.05 μA/cm 2 in WT, and 1.04 + 0.07 μA/cm 2 in Furin Mut, p < 0.05). In parallel, the expression of α-ENaC but not β or γ subunit as assessed by immunoblotting and qRT-PCR analysis was elevated by WT PRR and this increase was blunted by Furin Mut. In a separate experiment, M1 cells were transfected with pcDNA3.4 containing cDNA for sPRR with S1P cleavage (AA 1-282) (sPRR-S1P) or with furin cleavage (AA 1-279) (sPRR-furin); empty vector was used as a control. Overexpression of cDNA for the two types of sPRR induced a significant and comparable increase in the release of sPRR. By Ussing chamber technique, ENaC activity was 1.00 + 0.09 μA/cm 2 in EM, 1.03 + 0.10 μA/cm 2 in sPRR-S1P, and 1.39 + 0.14 μA/cm 2 in sPRR-furin, p < 0.05. Lastly, 293 cells were pretreated for 1 h with furin inhibitor α1-antitrypsin Portland followed by transfection with empty vector, WT PRR, or Furin Mut vectors. sPRR in the condition medium was enriched by using protein centrifugal filter devices and applied to M1 cells for 10 min followed by measurement of ENaC activity. Pretreatment with furin inhibition attenuated ENaC-acting activity induced by overexpression of WT PRR. Overall, the three independent approaches consistently demonstrated that furin-mediated modification is required for the activity of sPRR to increase ENaC-mediated Na + transport in the CD cells.

Effect of ammonia on Na+ transport across isolated rumen epithelium of sheep is diet dependent

2003 ◽  
Vol 90 (4) ◽  
pp. 751-758 ◽  
Author(s):  
Khalid Abdoun ◽  
Katarina Wolf ◽  
Gisela Arndt ◽  
Holger Martens
Keyword(s):  
Cellular Uptake ◽  
Ussing Chamber ◽  
Ammonium Ion ◽  
Na Transport ◽  
Adaptation Mechanism ◽  
Rumen Epithelium ◽  
Ruminal Epithelium ◽  
Positive Effect ◽  
Chamber Technique

The cellular uptake of ammonia affects the intracellular pH (pHi) of polar and non-polar cells. A predominant uptake of NH3 and its intra-cellular protonation tend to alkalinise the cytoplasm, whereas a predominant uptake of NH4+ acidifies the cytoplasm by reversing this reaction. Hence, the well-known absorption of ammonia across the rumen epithelium probably causes a change in the pHi. The magnitude and direction of this change in pHi (acid or alkaline) depends on the relative transport rates of NH3 and NH4+. Consequently, the intra-cellular availability of protons will influence the activity of the Na+-H+ exchanger, which could affect transepithelial Na+ transport. The aim of the present study has been to test this possible interaction between ruminal ammonia concentrations and Na+ transport. The term ammonia is used to designate the sum of the protonated (NH4+) and unprotonated (NH3) forms. Isolated ruminal epithelium of sheep was investigated by using the Ussing-chamber technique in vitro. The present results indicate that ammonia inhibits Na+ transport across the rumen epithelium of hay-fed sheep, probably by binding intracellular protons and thus inhibiting Na+-H+ exchange. By contrast, ammonia stimulates Na+ transport in concentrate-fed and urea-fed sheep, which develop an adaptation mechanism in the form of an increased metabolism of ammonia in the rumen mucosa and/or an increased permeability of rumen epithelium to the charged ammonium ion (NH4+). Intracellular dissociation of NH4+ increases the availability of protons, which stimulate Na+ –H+ exchange. This positive effect of ruminal ammonia on Na+ absorption may significantly contribute to the regulation of osmotic pressure of the ruminal fluid, because intraruminal ammonia concentrations up to 40 mmol/l have been reported.

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