Sugar uptake by intestinal basolateral membrane vesicles

Abstract

A high yield of membrane vesicles was prepared from the basolateral surface of rat intestinal cells using an N 2 cavitation bomb and density gradient centrifugation. The membranes were enriched 10-fold and were free of significant contamination by brush border membranes and mitochondria. The rate of d-[ 14C]-glucose and l-[ 3H]glucose uptake into the vesicles was measured using a rapid filtration technique. d-Glucose equilibrated within the vesicles with a half-time 1/25th that for l-glucose. The stereospecific uptake exhibited saturation kinetics with a K m of approx. 44 mM and a V of approx. 110 nmol · mg −1 · min −1 at 10°C. The activation energy for the process was 14 kcal · mol −1 below 15°C and it approached 3 kcal · mol −1 above 22°C. Carrier-mediated uptake was eliminated in the presence of 1 mM HgCl 2 and 0.5 mM phloretin. The rate of transport was unaffected by the absence or presence of sodium concentration gradients. Competition studies demonstrated that all sugars with the d-glucose pyranose ring chair conformation shared the transport system, and that, with the possible exception of the -OH group at carbon No. 1, there were no specific requirements for an equatorial -OH group at any position in the pyranose ring. In the case of α-methyl- d-glucoside its inability to share the d-glucose transport system may be due to steric hindrance posed by the -OCH 3 group rather than by a specific requirement for a free hydroxyl group at this position in the ring. It is concluded that sugars are transported across the basolateral membrane of the intestinal epithelium by a facilitated diffusion system reminiscent of that in human red blood cells.