Compromised white matter integrity in obesity.

Obesity is associated with both structural and functional changes of the central nervous system. While gray matter alterations in obesity point to a consistent reduction with increasing body mass index (BMI), volumetric changes in white matter are more complex and less conclusive. Hence, more recently, diffusion tensor imaging (DTI) has been employed as a highly sensitive tool to investigate microstructural changes in white matter structure. Parameters of diffusivity and anisotropy are used to evaluate white matter and fibre integrity as well as axonal and myelin degeneration. Fractional anisotropy (FA) is the most commonly used parameter as it is the best estimate of fibre integrity. The focus of this review was on the relationship between obesity and brain alterations assessed by DTI. Altogether, these studies have shown a loss of white matter integrity with obesity-related factors, especially in tracts within the limbic system and those connecting the temporal and frontal lobe. More specifically, multiple studies found an inverse association between BMI and FA in the corpus callosum, fornix, cingulum and corona radiata in elderly and young adults as well as children. Furthermore, significant interactions were observed between BMI and age, pointing to accelerated ageing of white matter structure in obese.

Involvement of the sodium-calcium exchanger 3 (NCX3) in ziram-induced calcium dysregulation and toxicity.

Ziram is a dimethyldithiocarbamate fungicide which can cause intraneuronal calcium (Ca(2+)) dysregulation and subsequently neuronal death. The signaling mechanisms underlying ziram-induced Ca(2+) dyshomeostasis and neurotoxicity are not fully understood. NCX3 is the third isoform of the sodium-calcium exchanger (NCX) family and plays an important role in regulating Ca(2+) homeostasis in excitable cells. We previously generated a mouse model deficient for the sodium-calcium exchanger 3 and showed that NCX3 is protective against ischemic damage. In the present study, we aim to examine whether NCX3 exerts a similar role against toxicological injury caused by the pesticide ziram. Our data show baby hamster kidney (BHK) cells stably transfected with NCX3 (BHK-NCX3) are more susceptible to ziram toxicity than cells transfected with the empty vector (BHK-WT). Increased toxicity in BHK-NCX3 was associated with a rapid rise in cytosolic Ca(2+) concentration [Ca(2+)]i induced by ziram. Profound mitochondrial dysfunction and ATP depletion were also observed in BHK-NCX3 cells following treatment with ziram. Lastly, primary dopaminergic neurons lacking NCX3 (NCX3(-/-)) were less sensitive to ziram neurotoxicity than wildtype control dopaminergic neurons. These results demonstrate that NCX3 genetic deletion protects against ziram-induced neurotoxicity and suggest NCX3 and its downstream molecular pathways as key factors involved in ziram toxicity. Our study identifies new molecular events through which pesticides (e.g. ziram) can lead to pathological features of degenerative diseases such as Parkinson's disease and indicates new targets to slow down neuronal degeneration.

[The functional sport shoe parameter "torsion" within running shoe research--a literature review]. / Der funktionelle Sportschuhparameter "Torsion" in der Laufschuhforschung - ein Literaturüberblick.

Within the sport shoe area torsion is described as the twisting and decoupling of the rear-, mid- and forefoot along the longitudinal axis of the foot. Studies have shown that running shoes restrict the torsion of the foot and thus they increase the pronation of the foot. Based on the findings, it is recommended to design running shoes, which allow the natural freedom of movement of the foot. The market introduction of the first torsion concept through adidas(R) took place in 1989. Independently of the first market introduction, only one epidemiological study was conducted in the running shoe area. The study should investigate the occurrence of Achilles tendon problems of the athletes running in the new "adidas Torsion(R) shoes". However, further studies quantifying the optimal region of torsionability concerning the reduction of injury incidence are still missing. Newer studies reveal that the criterion torsion only plays a secondary roll regarding the buying decision. Moreover, athletes are not able to perceive torsionability as a discrete functional parameter. It is to register, that several workgroups are dealing intensively with the detailed analysis of the foot movement based on kinematic multi-segment-models. However, scientific as well as popular scientific contributions display that the original idea of the torsion concept is still not completely understood. Hence, the "inverse" characteristic is postulated. The present literature review leads to the deduction that the functional characteristics of the torsion concept are not fully implemented within the running shoe area. This implies the necessity of scientific studies, which investigate the relevance of a functional torsion concept regarding injury prevention based on basic and applied research. Besides, biomechanical studies should analyse systematically the mechanism and the effects of torsion relevant technologies and systems.

Two types of neurotransmitter release patterns in isolectin B4-positive and negative trigeminal ganglion neurons.

Mammalian nociceptors have been classified into subclasses based on differential neurotrophin sensitivity and binding of the plant isolectin B4 (IB4). Most of the nerve growth factor-responsive IB4-negative (IB4 (-)) nociceptors contain neuropeptides such as substance P and calcitonin gene-related peptide, whereas the glial-derived neurotrophic factor-responsive IB4-positive (IB4 (+)) neurons predominantly lack such neuropeptides. We hypothesized that the differences in neuropeptide content between IB4 (+) and (-) neurons might be reflected in differences in stimulated exocytosis and/or endocytosis. To address this, we monitored the secretory activity of acutely dissociated neurons from adult rat trigeminal ganglia (TRG) using cell membrane capacitance (Cm) measurements and the fluorescent membrane-uptake marker N-(3-triethylammoniumpropyl)-4-(6-(4-(diethylamino)phenyl)hexatrienyl)pyridinium dibromide (FM4-64). Cm measurements were performed under whole-cell voltage clamp and neurons were depolarized from -75 mV to +10 mV to elicit exocytosis. Both types of TRG neurons showed similarly-sized, calcium-dependent increases in Cm, demonstrating that both IB4 (+) and (-) TRG neurons are capable of stimulated exocytosis. However, the peak Cm of IB4 (+) neurons decayed faster toward baseline than that of IB4 (-) neurons. Also, IB4 (+) neurons had stable Cm responses to repeated stimuli whereas IB4 (-) neurons loss their secretory response during repeated stimulation. These data suggested that the IB4 (+) neurons possess a faster rate of endocytosis and vesicle replenishment than IB4 (-) neurons. To test this, we measured vesicle trafficking with the fluorescent membrane dye FM4-64. FM4-64 staining showed that IB4 (-) neurons exhibit a larger pool of endocytosed vesicles than IB4 (+) neurons because the peak fluorescence increases in IB4 (-) neurons were larger but slower than in IB4 (+) neurons. However, the recycled vesicles were released faster in IB4 (+) compared with IB4 (-) neurons. Taken together these data suggest that the IB4 (+) TRG neurons have faster exocytosis and endocytosis than the IB4 (-) neurons.

Chain extension of sugar delta-lactones with the enolate of tert-butyl bromoacetate and elaboration into functionalized C-ketosides, C-glycosides, and C-glucosyl glycines.

[structure: see text] We describe the synthesis of a series of exocyclic sugar epoxides 1 prepared in a one-step procedure from sugar delta-lactones with the enolate of tert-butyl bromoacetate. Ring opening of the sugar oxiranes provides C-ketosides while reduction affords functionalized C-glycosides bearing an alpha-hydroxy ester moiety. The alpha-hydroxy ester can be converted into C-glucosyl glycine analogues 2.

Proteins involved in synaptic vesicle trafficking.

Neurotransmitter release relies on a series of synaptic vesicle trafficking reactions. We have determined the molecular basis of these reactions by microinjecting, into 'giant' nerve terminals of squid, probes that interfere with presynaptic proteins. These probes affect neurotransmitter release and disrupt nerve terminal structure. From the nature of these lesions, it is possible to deduce the roles of individual proteins in specific vesicle trafficking reactions. This approach has revealed the function of more than a dozen presynaptic proteins and we hypothesize that neurotransmitter release requires the coordinated action of perhaps 50-100 proteins.

Combinatorial synthesis of carbohydrates.

Combinatorial chemistry principles have been applied to the generation of oligosaccharide libraries, both in solution and on the solid phase, with a view to producing inhibitors of carbohydrate-protein binding. The rich stereochemistry and high degree of functionalization of sugars has also resulted in their increasing use in the synthesis of glycomimetics and as scaffolds for the presentation of pharmacophore groupings to receptors that are noncarbohydrate-recognizing proteins.

Protein interactions implicated in neurotransmitter release.

Biochemical evidence indicates that the exocytotic release of neurotransmitters involves both evolutionary conserved membrane proteins, the SNAREs, as well as ubiquitous cytosolic fusion proteins, NSF and SNAPs. We have analyzed the biochemical properties and the physiological effects of these proteins. Our data suggest models how NSF, SNAPs and SNAREs may function in neurotransmitter exocytosis.

Interaction between the pili of Pseudomonas aeruginosa PAK and its carbohydrate receptor beta-D-GalNAc(1-->4)beta-D-Gal analogs.

Pseudomonas aeruginosa employs pili to mediate adherence to epithelial cell surface receptors. Previously, it has been shown that the pilus adhesin of P. aeruginosa PAK binds to the ganglioside asialo-GM1. In particular, it was found that the carbohydrate sequence beta-D-GalNAc(1-->4)beta-D-Gal is the minimal carbohydrate receptor sequence of asialo-GM1. To study the binding specificity of P. aeruginosa, O-modified and N-modified sugar analogs, where each hydroxyl group was substituted either by O-methyl or O-propyl and the acetamido group was changed to a propionamido group, were synthesized. The sugar analogs were evaluated as inhibitors in a competitive solid phase binding assay. The results demonstrate that the pili of P. aeruginosa PAK accepts a variety of sugar analogs possessing the sequence beta-D-GalNAc(1-->4)beta-D-Gal. Most sugar analogs bind with a similar order of magnitude (50% inhibitory concentration (IC50) = 60-130 microM) except for the 2-O-propyl derivative 7 (IC50 = 8 +/- 4 microM) compared with an IC50 of 79 +/- 18 microM for the native compound. The significant increase in binding affinity of 2-O-propyl derivative 7 suggests that improved inhibitors of adhesion may be prepared by introducing a hydrophobic side chain at the 2-position of galactose.

Regulation of neurotransmitter release kinetics by NSF.

NSF (N-ethylmaleimide-sensitive factor) is an adenosine triphosphatase (ATPase) that contributes to a protein complex essential for membrane fusion. The synaptic function of this protein was investigated by injecting, into the giant presynaptic terminal of squid, peptides that inhibit the ATPase activity of NSF stimulated by the soluble NSF attachment protein (SNAP). These peptides reduced the amount and slowed the kinetics of neurotransmitter release as a result of actions that required vesicle turnover and occurred at a step subsequent to vesicle docking. These results define NSF as an essential participant in synaptic vesicle exocytosis that regulates the kinetics of neurotransmitter release and, thereby, the integrative properties of synapses.

Two sites of action for synapsin domain E in regulating neurotransmitter release.

Synapsins, a family of synaptic vesicle proteins, have been shown to regulate neurotransmitter release; the mechanism(s) by which they act are not fully understood. Here we have studied the role of domain E of synapsins in neurotransmitter release at the squid giant synapse. Two squid synapsin isoforms were cloned and found to contain a carboxy (C)-terminal domain homologous to domain E of the vertebrate a-type synapsin isoforms. Presynaptic injection of a peptide fragment of domain E greatly reduced the number of synaptic vesicles in the periphery of the active zone, and increased the rate and extent of synaptic depression, suggesting that domain E is essential for synapsins to regulate a reserve pool of synaptic vesicles. Domain E peptide had no effect on the number of docked synaptic vesicles, yet reversibly inhibited and slowed the kinetics of neurotransmitter release, indicating a second role for synapsins that is more intimately associated with the release process itself. Thus, synapsin domain E is involved in at least two distinct reactions that are crucial for exocytosis in presynaptic terminals.

Interaction of the receptor binding domains of Pseudomonas aeruginosa pili strains PAK, PAO, KB7 and P1 to a cross-reactive antibody and receptor analog: implications for synthetic vaccine design.

The four synthetic peptide antigens, PAK 128-144, PAO 128-144, KB7 128-144 and P1 126-148, correspond in amino acid sequence to the C-terminal receptor binding regions of four strains (PAK, PAO, KB7, P1) of Pseudomonas aeruginosa pilin. The NMR solution structures of the trans forms of the peptides show conserved beta-turns which have been implicated in antibody and receptor recognition. The interactions between these peptides and a cross-reactive monoclonal antibody, PAK-13, have been studied using two-dimensional (1)H NMR spectroscopy in order to map the antigenic determinants recognized by the antibody. Residues for which spectral changes were observed upon antibody binding differed from peptide to peptide but were mostly confined to one or both of the turn regions and to the hydrophobic pockets. Conformational changes in the beta-turns and hydrophobic pockets of these peptides upon antibody binding were also monitored by examination of the pattern of nuclear Overhauser effects (NOEs) versus transferred nuclear Overhauser effects (TRNOEs) for the free versus the bound peptides. Although TRNOEs developed strongly between side chain resonances in the hydrophobic pockets of the peptides, no additional backbone TRNOEs were observed in the presence of antibody, suggesting no major conformational changes in the secondary structures of the peptides upon binding. This implies a flexible antibody combining site, a feature which is discussed with respect to cross-reactivity, strain specificity, and the design of a synthetic peptide vaccine effective against a broad spectrum of P. aeruginosa strains. The binding of the PAK peptide to a disaccharide receptor analog, (beta GalNAc(1-4)beta Gal), was also studied using (1)H NMR in order to map the "adhesintope" recognized by the receptor. Spectral changes observed in the peptide spectrum with the binding of receptor were similar to those seen for the binding of antibody, suggesting that the epitope recognized by the antibody is structurally coincident with the adhesintope recognized by the receptor. The relevancy of this result is discussed with respect to immunogenicity versus pathogenicity, and the proper design of a vaccine which could prevent the mutational escape of the pathogen away from the host's defence systems.

Experimental vitreous replacement with perfluorophenanthrene.

PURPOSE: We studied the toxicity of perfluorophenanthrene (PFP), used as short-, medium-, or long-term internal tamponading substance, on the rabbit choroid and retina. The aim was the check whether PFP is tolerated as a tamponade in the eye or damages the retina, and if any such damage is due to toxicity or to emulsification which may stimulate phagocytosis. METHODS: Twenty-four right eyes of rabbits operated by vitrectomy were filled with 0.8-1 ml of PFP as retinal tamponading substance. Twelve eyes were operated by vitrectomy and filled with 0.8-1 ml of balanced salt solution as the control group. Direct and indirect ophthalmoscopy was done on the third and seventh day after surgery, then once a week. Enucleation, with histological and immunohistochemical examination, was done on the second day, then in the first, second, fourth and eighth weeks after surgery. RESULTS AND CONCLUSIONS: Histological examination showed progressive damage of the chorioretinal tissues right from the second week: the chorioretinal structure was completely altered from the eighth week. Immunohistochemical examinations showed that glia cells were involved in the inflammation consequent to internal tamponade with PFP.

Exocytosis: proteins and perturbations.

Exocytosis is the primary means of cellular secretion. Because exocytosis involves fusion between the plasma membrane and the membrane of secretory vesicles, it is likely that proteins on these two membranes, as well as additional proteins in cellular cytoplasm, mediate exocytosis. Although we know much about the proteins of secretory cells, we still have much to learn about how these proteins participate in exocytosis; in no case has an unambiguous exocytotic function been assigned to any of these proteins. To identify the roles of proteins in exocytosis it is necessary to perturb protein function in living secretory cells. We review a number of perturbation strategies and summarize what this approach has taught us about the functional roles of proteins in exocytosis, concluding with a molecular model of protein dynamics during exocytosis.

SNAP-mediated protein-protein interactions essential for neurotransmitter release.

The constitutive fusion of transport vesicles with intracellular membranes requires soluble proteins called SNAPs. Certain presynaptic proteins implicated in synaptic vesicle exocytosis also bind SNAPs, suggesting that SNAPs participate in the calcium-regulated membrane fusion events mediating neurotransmitter release. Here we show that injection of recombinant SNAPs into the giant synapse of squid enhances transmitter release. Conversely, injection of peptides designed to mimic the sites at which SNAP interacts with its binding partners inhibits transmitter release downstream of synaptic vesicle docking. A SNAP-dependent protein complex must therefore mediate transmitter release, showing that transmitter release shares a common molecular mechanism with constitutive membrane fusion.

Nitric oxide synthase expression in single hippocampal neurons.

The presence of nitric oxide synthase (NOS) in CA1 pyramidal cells of the rat hippocampus was demonstrated by single-cell PCR. NOS-specific primers were used to amplify mRNA isolated from single hippocampal neurons. The sequence of the major amplification-product obtained was identical to that of the constitutively expressed brain-isoform of NOS. These results confirm immunocytochemical data that NOS is present in CA1, and, therefore, nitric oxide could function as a retrograde messenger in long-term potentiation.