The Effect of Vanadate on Glucase Transport of Everted Sacs and Vesicles Prepared from Small Intestinal Epithelial Cells

Vanadate has been reported to inhibit Na⁺, K⁺-ATPase of many cells. Since intestinal uptake of glucose is influenced by the enzyme, we studied the effects of varying concentrations of sodium vanadate(NaVO₃) on the glucose transport in everted sacs prepared from the segment of jejunum and upper part of ileum of rabbit.

At concentrations of range 10^-8M~10^-4M vanadate decreased the mucosal-to-serosal flux(S/M ratio) of glucose to 4.8% -38.8% accompanying inhibition of Na⁺, K⁺-ATPase activities of basolateral membranes and alkaline phosphatase activities of brush-border membranes purified from small intestinal epithelial cells.

On the other hand, it was found that there were two cytochalasin B binding sites at the basolateral membranes, of which site I showed a dissociation constant of 1.8×10^-8M with maximal binding capacity of 26 pmoles/mg protein and site II showed a dissociation constant of 9.3×10^-7M with maximal binding capacity of 140 pmoles/mg protein. Vanadate(10^-4M) inhibits cytochalasin B binding at the both binding sites to 38.5% and 45.7% respectively.

Vanadate also inhibited cytochalasin B binding at both ghost membranes and band 4.5(glucose carrier) protein purified from human red blood cell membranes.

SDS-polyacrylamide gel electrophoresis(SDS-PAGE) showed that there existed a band 4.5 like protein in both basolateral membrane and brush-border membrane.

Based on the above results the following conclusions are obtained:

1) Glucose is absorbed across the small intestinal epithelium by a two stage process. the first is sodium co-transport system of the brush-border membrane and the second is sodium-independent facilitated diffusion system of the basolateral membrane.

2) The driving force for the transcellular movement of glucose is mainly dependent on sodium concentration gradient across the brush-border membrane which is generated by Na⁺, K⁺ ATPase at the basolateral membrane and also dependent directly on the activity of glucose carrier which may exist at the basolateral membrane.

3) Vanadate inhibits glucose uptake across the intestinal epithelium by affecting on both the Na⁺, K⁺-ATPase and glucose carrier of the basolateral membrane.