Intracellular hypertonicity is responsible for water flux associated with Na+/glucose cotransport.
Detection of a significant transmembrane water flux immediately after cotransporter stimulation is the experimental basis for the controversial hypothesis of secondary active water transport involving a proposed stoichiometry for the human Na(+)/glucose cotransporter (SGLT1) of two Na(+), one glucose, and 264 water molecules. Volumetric measurements of Xenopus laevis oocytes coexpressing human SGLT1 and aquaporin can be used to detect osmotic gradients with high sensitivity. Adding 2 mM of the substrate alpha-methyl-glucose (alphaMG) created mild extracellular hypertonicity and generated a large cotransport current with minimal cell volume changes. After 20, 40, and 60 s of cotransport, the return to sugar-free, isotonic conditions was accompanied by measurable cell swelling averaging 0.051, 0.061, and 0.077 nl/s, respectively. These water fluxes are consistent with internal hypertonicities of 1.5, 1.7, and 2.2 mOsm for these cotransport periods. In the absence of aquaporin, the measured hypertonicites were 4.6, 5.0, and 5.3 mOsm for the same cotransport periods Cotransport-dependent water fluxes, previously assumed to be water cotransport, could be largely explained by hypertonicities of such amplitudes. Using intracellular Na(+) injection and Na(+)-selective electrode, the intracellular diffusion coefficient for Na(+) was estimated at 0.29 +/- 0.03 x 10(-5) cm(2) s(-1). Using the effect of intracellular alphaMG injection on the SGLT1-mediated outward current, the intracellular diffusion coefficient of alphaMG was estimated at 0.15 +/- 0.01 x 10(-5) cm(2) s(-1). Although these intracellular diffusion coefficients are much lower than in free aqueous solution, a diffusion model for a single solute in an oocyte would require a diffusion coefficient three times lower than estimated to explain the local osmolyte accumulation that was experimentally detected. This suggests that either the diffusion coefficients were overestimated, possibly due to the presence of convection, or the diffusion in cytosol of an oocyte is more complex than depicted by a simple model.
Charron
François M
FM
Groupe d'Etude des Protéines Membranaires, Département de Physique, Université de Montréal, Québec H3C 3J7, Canada.
Blanchard
Maxime G
MG
Lapointe
Jean-Yves
JY
D000818:Animals; D048429:Cell Size; D002478:Cells, Cultured; D004058:Diffusion; D005947:Glucose / Q000378:metabolism*; D007424:Intracellular Fluid / Q000378:metabolism; D009865:Oocytes / Q000166:cytology* / Q000502:physiology*; D009997:Osmotic Pressure; D051269:Sodium-Glucose Transporter 1 / Q000378:metabolism*; D014867:Water / Q000378:metabolism*; D014882:Water-Electrolyte Balance / Q000502:physiology*; D014982:Xenopus laevis
C495674:SLC5A1 protein, human; D051269:Sodium-Glucose Transporter 1; D014867:Water; D005947:Glucose