We have investigated the mechanism responsible for the diffusive component of intestinal glucose absorption, the major route by which glucose is absorbed. In perfused rat jejunum in vivo, absorption was strongly inhibited by phloretin, an inhibitor of GLUT2. The GLUT2 level at the brush-border membrane increased some 2-fold when the luminal glucose concentration was changed from 0 to 100 mM. The phloretin-sensitive or diffusive component of absorption appeared superficially linear and consistent with simple diffusion, but was in fact carrier-mediated and co-operative (n=1.6, [G(1/2)]=56 mM; where [G(1/2)] is the glucose concentration at half V(max)) because of the glucose-induced activation and recruitment of GLUT2 to the brush-border membrane. Diffusive transport by paracellular flow was negligible. The phloretin-insensitive, SGLT1-mediated, component of glucose absorption showed simple saturation kinetics with [G(1/2)]=27 mM: the activation of protein kinase C (PKC) betaII, the isoenzyme of PKC that most probably controls GLUT2 trafficking [Helliwell, Richardson, Affleck and Kellett (2000) Biochem. J. 350, 149-154], also showed simple saturation kinetics, with [G(1/2)]=21 mM. We conclude that the principal route for glucose absorption is by GLUT2-mediated facilitated diffusion across the brush-border membrane, which is up to 3-fold greater than that by SGLT1; the magnitude of the diffusive component at any given glucose concentration correlates with the SGLT1-dependent activation of PKC betaII. The implications of these findings for the assimilation of sugars immediately after a meal are discussed.

Perfusion of rat jejunum in vitro with PMA increased fructose transport by 70% compared with control values and was blocked by the protein kinase C (PKC) inhibitor chelerythrine. The brush-border membrane contained both the fructose transporters GLUT5 and GLUT2; the presence of the latter was confirmed by luminal biotinylation. PMA increased the GLUT2 level 4-fold within minutes, so that the level was comparable with that of the basolateral membrane, but had no effect on GLUT5 level. GLUT2 was functional, accessible to luminal fructose and could be inhibited selectively by phloretin to permit determination of GLUT2- and GLUT5-mediated transport components. The 4-fold increase in GLUT2 level induced by PMA was matched by a 4-fold increase in GLUT2-mediated transport: there was a compensatory fall in the GLUT5-mediated rate. The pattern of dynamic trafficking was seen only for GLUT2, not GLUT5 or SGLT1, implying that GLUT2 trafficks to the brush-border membrane by a different pathway. Trafficking of GLUT2 to the brush-border membrane correlated with activation of PKC betaII, implying that this isoenzyme is likely to control trafficking. Since PKC is activated by endogenous hormones, GLUT2 levels in vivo are 3-4-fold those in vitro; moreover, because PKC is inactivated as soon as intestine is excised, GLUT2 is lost from the brush-border within minutes in vitro. It is therefore difficult to detect GLUT2 in most in vitro preparations and its role in intestinal sugar absorption across the brush-border membrane has accordingly been overlooked.

T1R taste receptors are present throughout the gastrointestinal tract. Glucose absorption comprises active absorption via SGLT1 and facilitated absorption via GLUT2 in the apical membrane. Trafficking of apical GLUT2 is rapidly up-regulated by glucose and artificial sweeteners, which act through T1R2 + T1R3/α-gustducin to activate PLC β2 and PKC βII. We therefore investigated whether non-sugar nutrients are regulated by taste receptors using perfused rat jejunum in vivo. Under different conditions, we observed a Ca 2+ -dependent reciprocal relationship between the H + /oligopeptide transporter PepT1 and apical GLUT2, reflecting the fact that trafficking of PepT1 and GLUT2 to the apical membrane is inhibited and activated by PKC βII, respectively. Addition of l-glutamate or sucralose to a perfusate containing low glucose (20 mm) each activated PKC βII and decreased apical PepT1 levels and absorption of the hydrolysis-resistant dipeptide l-Phe( S)-l-Ala (1 mm), while increasing apical GLUT2 and glucose absorption within minutes. Switching perfusion from mannitol to glucose (75 mm) exerted similar effects. l-Glutamate induced rapid GPCR internalization of T1R1, T1R3 and transducin, whereas sucralose internalized T1R2, T1R3 and α-gustducin. We conclude that l-glutamate acts via amino acid and glucose via sweet taste receptors to coordinate regulation of PepT1 and apical GLUT2 reciprocally through a common enterocytic pool of PKC βII. These data suggest the existence of a wider Ca 2+ and taste receptor-coordinated transport network incorporating other nutrients and/or other stimuli capable of activating PKC βII and additional transporters, such as the aspartate/glutamate transporter, EAAC1, whose level was doubled by l-glutamate. The network may control energy supply.

We have investigated the mechanism responsible for the diffusive component of intestinal glucose absorption, the major route by which glucose is absorbed. In perfused rat jejunum in vivo, absorption was strongly inhibited by phloretin, an inhibitor of GLUT2. The GLUT2 level at the brush-border membrane increased some 2-fold when the luminal glucose concentration was changed from 0 to 100 mM. The phloretin-sensitive or diffusive component of absorption appeared superficially linear and consistent with simple diffusion, but was in fact carrier-mediated and co-operative (n=1.6, [G(1/2)]=56 mM; where [G(1/2)] is the glucose concentration at half V(max)) because of the glucose-induced activation and recruitment of GLUT2 to the brush-border membrane. Diffusive transport by paracellular flow was negligible. The phloretin-insensitive, SGLT1-mediated, component of glucose absorption showed simple saturation kinetics with [G(1/2)]=27 mM: the activation of protein kinase C (PKC) betaII, the isoenzyme of PKC that most probably controls GLUT2 trafficking [Helliwell, Richardson, Affleck and Kellett (2000) Biochem. J. 350, 149-154], also showed simple saturation kinetics, with [G(1/2)]=21 mM. We conclude that the principal route for glucose absorption is by GLUT2-mediated facilitated diffusion across the brush-border membrane, which is up to 3-fold greater than that by SGLT1; the magnitude of the diffusive component at any given glucose concentration correlates with the SGLT1-dependent activation of PKC betaII. The implications of these findings for the assimilation of sugars immediately after a meal are discussed.

No abstract