On a three-dimensional templated model of GLUT1 (Protein Data Bank code 1SUK), a molecular recognition program, AUTODOCK 3, reveals nine hexose-binding clusters spanning the entire "hydrophilic" channel. Five of these cluster sites are within 3-5 Å of 10 glucose transporter deficiency syndrome missense mutations. Another three sites are within 8 Å of two other missense mutations. D-Glucose binds to five sites in the external channel opening, with increasing affinity toward the pore center and then passes via a narrow channel into an internal vestibule containing four lower affinity sites. An external site, not adjacent to any mutation, also binding phloretin but recognizing neither D-fructose nor L-glucose, may be the main threading site for glucose uptake. Glucose exit from human erythrocytes is inhibited by quercetin (K i ‫؍‬ 2.4 M) but not anionic quercetin-semiquinone. Quercetin influx is retarded by extracellular D-glucose (50 mM) but not by phloretin and accelerated by intracellular D-glucose. Quercetin docking sites are absent from the external opening but fill the entire pore center. In the inner vestibule, Glu 254 and Lys 256 hydrogen-bond quercetin (K i ≈ 10 M) but not quercetin-semiquinone. Consistent with the kinetics, this site also binds D-glucose, so quercetin displacement by glucose could accelerate quercetin influx, whereas quercetin binding here will competitively inhibit glucose efflux. ␤-D-Hexoses dock twice as frequently as their ␣-anomers to the 23 aromatic residues in the transport pathway, suggesting that endocyclic hexose hydrogens, as with maltosaccharides in maltoporins, form -bonds with aromatic rings and slide between sites instead of being translocated via a single alternating site.The glucose uniporter GLUT1 (SLC2A1), a member of the major facilitator superfamily of solute transporters, has to date not been crystallized, but its three-dimensional structure has been modeled by templating it to that of Lac Y permease and glycerol 3-phosphate antiporter (GlpT) from Escherichia coli (1-3). The 12 transmembrane ␣-helical domains of the monomeric GLUT protein are arranged around a central water-filled pore lined predominantly with uncharged hydrophilic and hydrophobic amino acids. The 15-Å-long, 8-Å-wide channel narrows near its midpoint (1, 2). Molecular dynamic simulations show that glucose binds close to this position within the pore, as expected of lactose binding to Lac Y permease (2). Glucose docks additionally in a cavity at the external entrance of the pore (3).GLUTs transport other substrates besides hexoses (e.g. dehydroascorbate (4, 5) via GLUT1, -3, and -4 and glucosamine (6) via GLUT2). The flavonone, quercetin, is transported via GLUT4. Quercetin influx into GLUT4 is inhibited by high glucose or cytochalasin B concentrations (7). Conversely, quercetin inhibits glucose and ascorbate transport via GLUT1, -2, -3, and -4 (8 -10).This present study demonstrates that quercetin is also transported via GLUT1, and its uptake is accelerated by exchange with intracellular glucose. Our...

Fluid and electrolyte absorption by colonic crypts depends on the transport properties of crypt cellular and paracellular routes and of the pericryptal sheath. As a low-Na(+) diet increases aldosterone and angiotensin II secretion, either hormone could affect absorption. Control and adrenalectomized (ADX) Sprague-Dawley rats were kept at a high-NaCl (HS) diet and then switched to low-NaCl (LS) diet for 3 days. Aldosterone or angiotensin II plasma concentrations were maintained using implanted osmotic mini-pumps. The extracellular Na(+) concentration in isolated rat distal colonic mucosa was determined by confocal microscopy using a low-affinity Na(+) -sensitive fluorescent dye (Sodium red, and Na(+) -insensitive BODIPY) bound to polystyrene beads. Crypt permeability to FITC-labelled dextran (10 kDa) was monitored by its rate of escape from the crypt lumen into the pericryptal space. Mucosal ion permeability was estimated by transepithelial electrical resistance (TER) and amiloride-sensitive short-circuit current (SCC). The epithelial Na(+) channel, ENaC, was determined by immunolocalization. LS diet decreased crypt wall permeability to dextran by 10-fold and doubled TER. Following ADX, aldosterone decreased crypt wall dextran permeability, increased TER, increased Na(+) accumulation in the pericryptal sheath and ENaC expression even in HS. Infusion of angiotensin II to ADX rats did not reverse the effects of aldosterone deprivation. These findings indicate that aldosterone alone is responsible for both the increase in Na(+) absorption and the decreased paracellular and pericryptal sheath permeability.

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Pericryptal myofibroblast growth in descending colonic crypts correlates with the activation of the renin-angiotensin-aldosterone system. Earlier work showed that during the transition from a high-Na(+) (HS) to low-Na(+) (LS) diet there are changes in the colonic crypt wall and pericryptal sheath. As LS diet increases both aldosterone and angiotensin II, the aim here was to determine their individual contributions to the trophic changes in colonic crypts. Experiments were conducted on control and adrenalectomized Sprague-Dawley rats fed an HS diet and then switched to LS diet for 3 days and supplemented with aldosterone or angiotensin II. The actions of the angiotensin-converting enzyme inhibitor captopril, the angiotensin receptor antagonist losartan and the aldosterone antagonist spironolactone on extracellular matrix proteins, claudin 4 and E-cadherin myofibroblast proteins, alpha-smooth muscle actin (alpha-SMA) and OB-cadherin (cadherin 11), angiotensin type 1 and TGFbetar1 membrane receptors were determined by immunolocalization in fixed distal colonic mucosa. The LS diet or aldosterone supplementation following ADX in HS or LS increased extracellular matrix, membrane receptors and myofibroblast proteins, but angiotensin alone had no trophic effect on alpha-SMA. These results show that aldosterone stimulates myofibroblast growth in the distal colon independently of dietary Na(+) intake and of angiotensin levels. This stimulus could be a genomic response or secondary to stretch of the pericryptal sheath myofibroblasts accompanying enhanced rates of crypt fluid absorption.

1 Nootropic drugs increase glucose uptake into anaesthetised brain and into Alzheimer's diseased brain. Thyrotropin-releasing hormone, TRH, which has a chemical structure similar to nootropics increases cerebellar uptake of glucose in murine rolling ataxia. This paper shows that nootropic drugs like piracetam (2-oxo 1 pyrrolidine acetamide) and levetiracetam and neuropeptides like TRH antagonise the inhibition of glucose transport by barbiturates, diazepam, melatonin and endogenous neuropeptide galanin in human erythrocytes in vitro. 2 The potencies of nootropic drugs in opposing scopolamine-induced memory loss correlate with their potencies in antagonising pentobarbital inhibition of erythrocyte glucose transport in vitro (Po0.01). Less potent nootropics, D-levetiracetam and D-pyroglutamate, have higher antagonist K i 's against pentobarbital inhibition of glucose transport than more potent L-stereoisomers (Po0.001).3 Piracetam and TRH have no direct effects on net glucose transport, but competitively antagonise hypnotic drug inhibition of glucose transport. Other nootropics, like aniracetam and levetiracetam, while antagonising pentobarbital action, also inhibit glucose transport. Analeptics like bemigride and methamphetamine are more potent inhibitors of glucose transport than antagonists of hypnotic action on glucose transport. 4 There are similarities between amino-acid sequences in human glucose transport protein isoform 1 (GLUT1) and the benzodiazepine-binding domains of GABA A (gamma amino butyric acid) receptor subunits. Mapped on a 3D template of GLUT1, these homologies suggest that the site of diazepam and piracetam interaction is a pocket outside the central hydrophilic pore region. 5 Nootropic pyrrolidone antagonism of hypnotic drug inhibition of glucose transport in vitro may be an analogue of TRH antagonism of galanin-induced narcosis .