Abstract
The cloned rabbit intestinal Na+/glucose cotransporter was expressed in Xenopus oocytes, and transmembrane currents associated with this transporter were monitored using a two-electrode voltage clamp. Addition of D-glucose to a Na(+)-containing solution bathing these oocytes generated a current which was blocked by phlorizin. Water-injected control oocytes did not exhibit any currents under these conditions. The magnitude and shape of the currents were dependent on the extracellular glucose and Na+ concentrations and the membrane potential. At Vhold = -50 mV, the Km values for glucose and Na+ were 14 +/- 2 (N = 4) microM and 17 +/- 1 (N = 3) mM, respectively. These Km values and imax exhibited voltage dependence: increasing the membrane potential from -30 to -150 mV increased KGlcm and imax threefold and decreased KNam eightfold. The reversal potential (VR) of the phlorizin-sensitive, glucose-dependent current varied with log Nao+ (slope 46 +/- 6 [N = 9] mV). In the absence of sugar, a Na(+)-dependent, phlorizin-sensitive (Ki = 3 +/- 0.5 microM) current was detected only in RNA-injected oocytes. The amplitude of this current at -50 mV was 6 +/- 1% (N = 13) of the maximum current measured in the presence of D-glucose. The VR of this sugar-independent current varied with log Nao+ (slope 63 +/- 1 [N = 4] mV), indicating that the cotransporter may carry Na+ in the absence of sugar. We conclude that the Na+/glucose cotransporter is electrogenic and that investigations of currents associated with its operation can yield valuable insights into the mechanisms of solute translocation.
MeSH Terms
Animals
Electric Conductivity
Female
In Vitro Techniques
Intestines/physiology
Ion Channels/drug effects,physiology
Kinetics
Mathematics
Membrane Potentials/drug effects
Methylglucosides/pharmacology
Microinjections
Models, Theoretical
Monosaccharide Transport Proteins/genetics,physiology
Oocytes/physiology
RNA, Messenger/administration & dosage,genetics
Rabbits
Regression Analysis
Sodium/pharmacology
Xenopus laevis
Chemicals
Ion Channels
Methylglucosides
Monosaccharide Transport Proteins
RNA, Messenger
methylglucoside
Sodium
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Umbach J A
Department of Pharmacology, Jerry Lewis Neuromuscular Research Center, Los Angeles, California.
Coady M J
Wright E M
References (16)
16 references, click to expand
-
METHOD FOR ASSAY OF INTESTINAL DISACCHARIDASES.
Anal Biochem. 1964 Jan;7:18-25
PMID: 14106916
-
Kinetic analysis of a family of cotransport models.
Biochim Biophys Acta. 1981 Dec 7;649(2):269-80
PMID: 7317398
-
The use of Xenopus oocytes for the study of ion channels.
CRC Crit Rev Biochem. 1987;22(4):317-87
PMID: 2449311
-
Expression cloning and cDNA sequencing of the Na+/glucose co-transporter.
Nature. 1987 Nov 26-Dec 2;330(6146):379-81
PMID: 2446136
-
Characterization of a Na+/glucose cotransporter cloned from rabbit small intestine.
J Membr Biol. 1989 Aug;110(1):87-95
PMID: 2795642
-
Na+-coupled sugar transport: membrane potential-dependent Km and Ki for Na+.
Am J Physiol. 1988 Oct;255(4 Pt 1):C486-94
PMID: 3177623
-
Sodium-induced conformational changes in the glucose transporter of intestinal brush borders.
J Biol Chem. 1984 Nov 25;259(22):14105-12
PMID: 6438091
-
Characterization of the D-glucose/Na+ cotransport system in the intestinal brush-border membrane by using the specific substrate, methyl alpha-D-glucopyranoside.
Biochim Biophys Acta. 1987 Nov 2;904(1):71-80
PMID: 3663668
-
Electrogenic properties of the sodium-alanine cotransporter in pancreatic acinar cells: I. Tight-seal whole-cell recordings.
J Membr Biol. 1986;94(2):99-115
PMID: 3560201
-
Electrogenic properties of the sodium-alanine cotransporter in pancreatic acinar cells: II. Comparison with transport models.
J Membr Biol. 1986;94(2):117-27
PMID: 3560198
-
Current-voltage relations of sodium-coupled sugar transport across the apical membrane of Necturus small intestine.
J Membr Biol. 1986;93(3):205-19
PMID: 3820278
-
The mechanistic nature of the membrane potential dependence of sodium-sugar cotransport in small intestine.
J Membr Biol. 1985;87(2):159-72
PMID: 4078884
-
Xenopus oocyte resting potential, muscarinic responses and the role of calcium and guanosine 3',5'-cyclic monophosphate.
J Physiol. 1984 Jul;352:551-74
PMID: 6086916
-
Kinetics of sodium-dependent solute transport by rabbit renal and jejunal brush-border vesicles using a fluorescent dye.
J Physiol. 1983 Feb;335:307-18
PMID: 6875880
-
Kinetics of sodium D-glucose cotransport in bovine intestinal brush border vesicles.
J Membr Biol. 1984;79(1):41-51
PMID: 6737463
-
Expression of an omega-conotoxin-sensitive calcium channel in Xenopus oocytes injected with mRNA from Torpedo electric lobe.
Proc Natl Acad Sci U S A. 1987 Aug;84(15):5464-8
PMID: 2440049