Abstract:The lessons learned from p-octiphenyl -barrel pores are applied to the rational design of synthetic multifunctional pore 1 that is unstable but inert, two characteristics proposed to be ideal for practical applications. Nonlinear dependence on monomer concentration provided direct evidence that pore 1 is tetrameric (n ) 4.0), unstable, and "invisible," i.e., incompatible with structural studies by conventional methods. The long lifetime of high-conductance single pores in planar bilayers demonstrated that rigid-rod -barrel 1 is inert and large (d ≈ 12 Å). Multifunctionality of rigid-rod -barrel 1 was confirmed by adaptable blockage of pore host 1 with representative guests in planar (8-hydroxy-1,3,6-pyrenetrisulfonate, KD ) 190 µM, n ) 4.9) and spherical bilayers (poly-L-glutamate, KD e 105 nM, n ) 1.0; adenosine triphosphate, KD ) 240 µM, n ) 2.0) and saturation kinetics for the esterolysis of a representative substrate (8-acetoxy-1,3,6-pyrenetrisulfonate, KM ) 0.6 µM). The thermodynamic instability of rigid-rod -barrel 1 provided unprecedented access to experimental evidence for supramolecular catalysis (n ) 3.7). Comparison of the obtained kcat ) 0.03 min -1 with the kcat ≈ 0.18 min -1 for stable analogues gave a global KD ≈ 39 µM 3 for supramolecular catalyst 1 with a monomer/barrel ratio ≈ 20 under experimental conditions. The demonstrated "invisibility" of supramolecular multifunctionality identified molecular modeling as an attractive method to secure otherwise elusive insights into structure. The first molecular mechanics modeling (MacroModel, MMFF94) of multifunctional rigid-rod -barrel pore hosts 1 with internal 1,3,6-pyrenetrisulfonate guests is reported.

The retina of honeybee drone is a nervous tissue with a crystal-like structure in which glial cells and photoreceptor neurons constitute two distinct metabolic compartments. The phosphorylation of glucose and its subsequent incorporation into glycogen occur in glia, whereas O2 consumption (QO2) occurs in the photoreceptors. Experimental evidence showed that glia phosphorylate glucose and supply the photoreceptors with metabolic substrates. We aimed to identify these transferred substrates. Using ion-exchange and reversed-phase HPLC and gas chromatography-mass spectrometry, we demonstrated that more than 50% of 14C(U)-glucose entering the glia is transformed to alanine by transamination of pyruvate with glutamate. In the absence of extracellular glucose, glycogen is used to make alanine; thus, its pool size in isolated retinas is maintained stable or even increased. Our model proposes that the formation of alanine occurs in the glia, thereby maintaining the redox potential of this cell and contributing to NH3 homeostasis. Alanine is released into the extracellular space and is then transported into photoreceptors using an Na(+)-dependent transport system. Purified suspensions of photoreceptors have similar alanine aminotransferase activity as glial cells and transform 14C- alanine to glutamate, aspartate, and CO2. Therefore, the alanine entering photoreceptors is transaminated to pyruvate, which in turn enters the Krebs cycle. Proline also supplies the Krebs cycle by making glutamate and, in turn, the intermediate alpha-ketoglutarate. Light stimulation caused a 200% increase of QO2 and a 50% decrease of proline and of glutamate. Also, the production of 14CO2 from 14C-proline was increased. The use of these amino acids would sustain about half of the light-induced delta QO2, the other half being sustained by glycogen via alanine formation. The use of proline meets a necessary anaplerotic function in the Krebs cycle, but implies high NH3 production. The results showed that alanine formation fixes NH3 at a rate exceeding glutamine formation. This is consistent with the rise of a glial pool of alanine upon photostimulation. In conclusion, the results strongly support a nutritive function for glia.

We report the characterization of multifunctional rigid-rod β-barrel ion channels with either internal aspartates or arginine-histidine dyads by planar bilayer conductance experiments. Barrels with internal aspartates form cation selective, large, unstable and ohmic barrel-stave (rather than toroidal) pores; addition of magnesium cations nearly deletes cation selectivity and increases single-channel stability. Barrels with internal arginine-histidine dyads form cation selective (P K ϩ/P Cl Ϫ = 2.1), small and ohmic ion channels with superb stability (single-channel lifetime > 20 seconds). Addition of "protons" results in inversion of anion/cation selectivity (P Cl Ϫ/P K ϩ = 3.8); addition of an anionic guest (HPTS) results in the blockage of anion selective but not cation selective channels. These results suggest that specific, internal counterion immobilization, here magnesium (but not sodium or potassium) cations by internal aspartates and inorganic phosphates by internal arginines (but not histidines), provides access to synthetic multifunctional pores with attractive properties.

The retina of the honeybee drone is a nervous tissue in which glial cells and photoreceptor cells (sensory neurons) constitute two distinct metabolic compartments. Retinal slices incubated with 2-deoxy[3lHjglucose convert this glucose analogue to 2-deoxy[3H~glucose 6-phosphate, but this conversion is made only in the glial cells. Hence, glycolysis occurs only in glial cells. In contrast, the neurons consume 02 and this consumption is sustained by the hydrolysis of glycogen, which is contained in large amounts in the glia. During photostimulation the increased oxidative metabolism of the neurons is sustained by a higher supply of carbohydrates from the glia. This clear case of metabolic interaction between neurons and glial cells supports Golgi's original hypothesis, proposed nearly 100 years ago, about the nutritive function of glial cells in the nervous system. The hypothesis that one ofthe functions of glia in the nervous system is to transfer nutrients from the capillary blood to neurons was proposed by Golgi nearly a century ago (1, 2). This hypothesis seemed reasonable based upon histological evidence, but direct experimental support has so far been either meager or even negative (3). A question, often appearing in textbooks but as yet not answered, that bears upon the truth of this hypothesis is whether glycogenolysis in the glia serves to nourish the neurons. The question arises because histological observations show that in many nervous systems glial cells contain much more glycogen than neurons do (4-6). In the mammalian brain the glycogen is labile and it can be almost depleted by, for example, 1 min of ischemia (7) or hypoxia (8). Hence, it is known that carbohydrate stored as glycogen can be mobilized, but it is not known whether it is transferred from glia to neurons rather than being consumed by the glia themselves (9).We have examined the nutritive role of glial cells in a comparatively simple nervous tissue, the retina of the honeybee (Apis mellifera) drone. In this preparation the separation of metabolic functions between the glial cells and photoreceptor cells (sensory neurons) is exceptionally complete. The photoreceptors contain large numbers of mitochondria and very little glycogen. In contrast, the glial cells contain very few mitochondria, but large quantities of glycogen ,3 particles (10-12). Photoreceptor energy metabolism is obligatorily aerobic, since anoxia and the mitochondrial inhibitor amobarbital rapidly abolish light-induced electrical activity in the drone retina (11,13 MATERIALS AND METHODS Preparation of Slices and Uptake of Labeled Hexoses.Retinal slices (-250 ,um thick) were prepared by making two parallel cuts with a vibrating razor blade, parallel to the ommatidia in the dorsal region of the retina, as described elsewhere (12). When such a slice is exposed to Ringer solution oxygenated with 100% 02, the retina is well oxygenated throughout its full thickness (14). The Ringer solution normally contained 270 mM NaCl, 10 mM KCl, 10 mM MgCl2, 1.6 mM CaC12 and 10 mM t...

The self-assembly of the first pentanuclear helicate was predicted on the structural basis obtained for linear and tetranuclear parent supramolecular compounds. Accordingly, the designed ternary supramolecular system requires appropriate polytopic organic receptors, which were successfully synthesized. Indeed, the formation of pentanuclear complexes was experimentally evidenced with NMR and ESMS spectra that perfectly reflect the expected pattern. The structural features in the europium pentanuclear complex are highlighted with semiempirical molecular modeling. The present work validates the combinatorial approach leading to the thermodynamically driven formation of tower-like pentanuclear edifices.