Applications of confocal microscopy to the study of myelin development and neuron structure

Confocal laser scanning microscopy has been used to study the localization of myelin basic proteins expressed in nonglial cells, and to probe the three-dimensional structure of central auditory neurons in the lateral superior olive. The paper focuses on the techniques used to obtain the results. The key roles of confocal microscopy and computer image processing of the images obtained are emphasized as they relate to the discovery of essential structural information about these specimens.     

Molecular basis of the constitutive activity and STI571 resistance of Asp816Val mutant KIT receptor tyrosine kinase

The receptor tyrosine kinase, KIT, displays activating mutations in the kinase domain, which are associated with various cancers. We have used homology modelling based on the crystal structures of the insulin receptor kinase in active and inactive conformations to predict the corresponding structures of the KIT kinase domain. We have prepared four KIT models, one each for the active and inactive conformations of the wild-type and of the Asp816Val mutant proteins. We have also placed ATP into the active conformations and the inhibitor, STI571, into the inactive conformations. All models have been fully energy minimised. The molecular modelling studies described here explain (i) why Asp816Val KIT is constitutively active, (ii) why the nature of the substituting amino acid at residue 816 is relatively unimportant, and (iii) why the Asp816Val substitution confers resistance to the KIT-inhibitory drug STI571. The models will be valuable for predicting other kinase inhibitory drugs that may be active on wild-type and mutant forms of KIT. During the course of this work, a crystal structure of the active conformation of the KIT kinase domain has been published. Our model of the active conformation of the Asp816Val mutant is strikingly similar to this crystal structure, whereas our model of the active conformation of the wild-type kinase domain of KIT differs from the crystal structure in some respects. The reasons for this apparent discrepancy are discussed.     

Isolation of feeding deterrents against argentine stem weevil from ryegrass infected with the endophyteAcremonium loliae

Infection of ryegrass (Lolium perenne) by an endophytic fungus (Acremonium loliae) confers resistance against the Argentine stem weevil (Listronotus bonariensis). Extracts from ryegrass clones, infected and uninfected withA. loliae, were compared in a feeding choice bioassay, and several fractions were identified which affected stem weevil feeding behavior. One stem weevil feeding deterrent, peramine C₁₂H₁₇N₅O, has been isolated from infected ryegrass and partially characterized as a basic indole derivative. Extracts from culturedA. loliae had no effect on stem weevil feeding behavior nor was peramine detected in the fungal cultures examined. Peramine and the other active substances are hydrophilic in contrast to the lipophilic properties reported for the neurotoxic lolitrems also isolated from ryegrass infected withA. loliae and associated with causing ryegrass staggers disorder in livestock. It is suggested that ryegrass staggers and stem weevil feeding deterrency may arise by different biochemical mechanisms.     

The role of oncostatin M in animal and human connective tissue collagen turnover and its localization within the rheumatoid joint

OBJECTIVE: To study the interaction of interleukin-1alpha (IL-1alpha) and oncostatin M (OSM) in promoting cartilage collagen destruction. METHODS: Bovine, porcine, and human cartilage and human chondrocytes were studied in culture. The levels of collagenase (matrix metalloproteinase 1 [MMP-1]) and tissue inhibitor of metalloproteinases 1 (TIMP-1) were measured by bioassay and enzyme-linked immunosorbent assay (ELISA). The levels of OSM in rheumatoid synovial fluid were measured by ELISA. RESULTS: When combined with OSM, IL-1alpha, IL-1beta, and tumor necrosis factor alpha released proteoglycan and collagen from cartilage. OSM was the only member of the IL-6 family to have this effect. Human tendon also responded to IL-1alpha and OSM. OSM increased the production of MMP-1 and TIMP-1 but when combined with IL-1alpha, synergistically promoted MMP-1 production in human chondrocytes and synovial fibroblasts. High levels of OSM were found in human rheumatoid synovial fluids, and confocal microscopy showed that OSM was produced by macrophages in rheumatoid synovial tissue. CONCLUSION: These results highlight an important new mechanism by which there is irreversible loss of collagen from cartilage.     

Carbonaceous nanomaterials for the enhancement of TiO2 photocatalysis

Semiconductor photocatalysis has important applications such as achieving sustainable energy generation and treating environmental pollution. TiO₂ has been the most widely-researched photocatalyst, but suffers from low efficiency and narrow light response range. Combining TiO₂ with carbonaceous nanomaterials is being increasingly investigated as a means to increase photocatalytic activity, and demonstrations of enhancement are plentiful. This review surveys the literature and highlights recent progress in the development of nanocarbon-TiO₂ photocatalysts, covering activated carbon, carbon doping, carbon nanotubes, [60]-fullerenes, graphene, thin layer carbon coating, nanometric carbon black and more recently developed morphologies. Mechanisms of enhancement, synthesis routes and future applications are summarised and discussed. New insight and enhanced photocatalytic activity may be provided by novel nanocarbon-TiO₂ systems. Ongoing challenges and possible new directions are outlined.     

Stimulation on the positive phase of hippocampal theta rhythm induces long-term potentiation that can Be depotentiated by stimulation on the negative phase in area CA1 in vivo

Long-term potentiation (LTP) of synaptic transmission induced by high-frequency stimulation (HFS) is considered to be a model for learning processes; however, standard HFS protocols consisting of long trains of HFS are very different from the patterns of spike firing in freely behaving animals. We have investigated the ability of brief bursts of HFS triggered at different phases of background theta rhythm to mimic more natural activity patterns. We show that a single burst of five pulses at 200 Hz given on the positive phase of tail pinch-triggered theta rhythm reliably induced LTP in the stratum radiatum of the hippocampus of urethane-anesthetized rats. Three of these bursts saturated LTP, and 10 bursts occluded the induction of LTP by long trains of HFS. Burst stimulation on the negative phase or at zero phase of theta did not induce LTP or long-term depression. In addition, stimulation with 10 bursts on the negative phase of theta reversed previously established LTP. The results show that the phase of sensory-evoked theta rhythm powerfully regulates the ability of brief HFS bursts to elicit either LTP or depotentiation of synaptic transmission. Furthermore, because complex spike activity of approximately five pulses on the positive phase of theta rhythm can be observed in freely moving rats, LTP induced by the present theta-triggered stimulation protocol might model putative synaptic plastic changes during learning more closely than standard HFS-induced LTP.     

Applications of electron paramagnetic resonance to studies of defects in insulators and semiconductors

Electron paramagnetic resonance (EPR) which is the absorption of electromagnetic waves in the microwave frequency domain has been used for many years to study magnetic dipoles in crystals subjected to magnetic fields. These magnetic dipoles arise from electrons in the material whose intrinsic spin is exposed due to processes such as doping with transition metal ions, doping with ions whose spin is not locked up with chemical bonding and irradiation with bond breaking energy. The sensitivity of the EPR technique depends on such experimental parameters as, incident microwave power, size of the sample, quality of the microwave cavity, number of paramagnetic spins participating in the absorption, width of the EPR resonance line, temperature of the sample and relaxation time of the spin system to the lattice. The experimental spectrum is described by parameters which by the application of basic quantum mechanics can be related to properties of the wave function of the electron spin in the crystal environment such as bonding directions to neighboring ions in the crystal lattice. Some examples of this analysis will be presented to show how EPR is used in the study of defects in Si and SiO₂.