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• Articles by S. Nussberger
• Articles by M.A. Hediger

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• On the Distribution of a Permeable Solute during Poiseuille ...

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• Glucose Accumulation Can Account for the Initial Water Flux ...

  Glucose Accumulation Can Account for the Initial Water Flux Triggered by Na/Glucose Cotransport Biophysical Journal, Volume 86, Issue 1, 1 January 2004, Pages 125-133 Marilène P. Gagnon, Pierre Bissonnette, Louis-Martin Deslandes, Bernadette Wallendorff and Jean-Yves Lapointe Abstract Over the last decade, several cotransport studies have led to the proposal of secondary active transport of water, challenging the dogma that all water transport is passive. The major observation leading to this interpretation was that a Na influx failed to reproduce the large and rapid cell swelling induced by Na/solute cotransport. We have investigated this phenomenon by comparing a Na/glucose (hSGLT1) induced water flux to water fluxes triggered either by a cationic inward current (using ROMK2K channels) or by a glucose influx (using GLUT2, a passive glucose transporter). These proteins were overexpressed in oocytes and assayed through volumetric measurements combined with double-electrode electrophysiology or radioactive uptake measurements. The osmotic gradients driving the observed water fluxes were estimated by comparison with the swelling induced by osmotic shocks of known amplitude. We found that, for equivalent cation or glucose uptakes, the combination of substrate accumulations observed with ROMK2 and GLUT2 are sufficient to provide the osmotic gradient necessary to account for a passive water flux through SGLT1. Despite the fact that the Na/glucose stoichiometry of SGLT1 is 2:1, glucose accumulation accounts for two-thirds of the osmotic gradient responsible for the water flux observed at =30s. It is concluded that the different accumulation processes for neutral versus charged solutes can quantitatively account for the fast water flux associated with Na/glucose cotransport activation without having to propose the presence of secondary active water transport. Abstract | Full Text | PDF (168 kb)

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Abstract

Transport of organic and inorganic solutes into and out of cells requires specialized transport proteins. Given a sufficiently sensitive analytical method for measuring cellular solute concentrations, it should be possible to monitor solute transport across the plasma membrane at the level of single cells. We report a capillary zone electrophoresis approach that is generally applicable to monitor solute transport into Xenopus laevis oocytes, requires only nanoliters of sample, and involves no radioactive materials. The sensitivity of capillary electrophoresis with UV detection is typically on the order of 10(-5)-10(-6) M, resulting in the mass detection limits in the low femtomole range. We show that capillary zone electrophoresis serves as a simple technique to measure solute transport into oocytes. Studies of the mammalian oligopeptide transporter PepT1 and the Na(+)- and K(+)-coupled epithelial and neuronal glutamate transporter EAAC1 expressed in oocytes demonstrate that transport of the dipeptide Trp-Gly via PepT1 and transport of Na+ and K+ via EAAC1 across the oocyte plasma membrane can be monitored by measuring intracellular tryptophan absorption and by indirect UV detection of inorganic ions, respectively. The CZE method allowed the simultaneous detection of changes of intracellular Na+ and K+ concentrations in response to EAAC1-mediated Na+ cotransport and K+ countertransport. This is the first report of a capillary zone electrophoresis-based quantitative analysis of intracellular components of a single cell in response to transport activity.