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Disrupting the brain to guide plasticity and improve behavior.

Neurones may be highly stable and nonplastic cellular structures, but they are engaged in dynamically changing, intrinsically plastic neural networks that provide a most energy efficient, spatially compact, and precise means to process input signals and generate adaptable responses to a changing environment. Neural plasticity is evolution's invention to enable the nervous system to escape the restrictions of its own genome (and its highly specialized cellular specification) and thus adapt to environmental pressures, physiologic changes, and experiences. At neural system level two steps of plasticity can be identified: unmasking existing connections that may be followed by establishment of new ones possibly even with integration of new neural structures and neurons. In any case, plastic changes may not necessarily represent a behavioral gain for a given subject, as they represent the mechanism for development and learning, as much as a cause of pathology and disease. The challenge is to learn enough about the mechanisms of plasticity to be able to guide them, suppressing changes that may lead to undesirable behaviors while accelerating or enhancing those that result in a behavioral benefit for the subject or patient. Neurostimulation, including noninvasive brain stimulation techniques, provide an opportunity to modulate brain plasticity in a controlled and specific manner. Such interventions to guide behavior or treat pathological symptomatology might be more immediate in their behavioral repercussion and thus more effective than approaches intent on addressing underlying genetic predispositions.

Animals↗

Physiology and anatomy of lenticel-like structures on leaves of Eucalyptus nitens and Eucalyptus globulus seedlings.

Intumescences or abnormal, non-pathogenic, blister-like protuberant growths, form on Eucalyptus globulus Labill. and, to a much lesser extent, Eucalyptus nitens (Deane and Maiden) Maiden leaves when plants are grown in a high relative humidity environment. We examined the histology of intumescences and their effects on leaf photosynthetic processes. Intumescences were induced by placing E. globulus and E. nitens seedlings in a relative humidity of 80% in a greenhouse for 5 days. Symptomatic and asymptomatic leaves of plants with intumescence development were compared with leaves of control plants. Light-saturated carbon dioxide (CO(2)) assimilation (A(max)) and responses of CO(2) assimilation (A) to varying intercellular CO(2) partial pressure (C(i)) were measured. Symptomatic and asymptomatic leaf samples were fixed and sectioned and cellular structure was examined. Intumescences greatly reduced the photosynthetic capacity of E. globulus leaves and were associated with reduced electron transport rate and ribulose bisphosphate (RuBP) regeneration capacity. Tissue necrotization and cellular collapse of the palisade mesophyll and deposition of phenolic compounds in the affected areas, probably reduced light penetration to photosynthesizing cells as well as reducing the amount of photosynthesizing tissue. Photosynthetic capacity of E. nitens was unaffected. The intumescences resembled simple lenticels, both morphologically and developmentally. To our knowledge, this is the first time that lenticel-like structures developed in response to environmental conditions have been described on leaves.

Carbon Dioxide↗

The emerging functions of endocannabinoid signaling during CNS development.

In the postnatal brain, endocannabinoids acting as retrograde messengers regulate the function of many synapses. By contrast, the understanding of endocannabinoid functions that regulate fundamental developmental processes such as cell proliferation, migration, differentiation and survival during patterning of the CNS is just beginning to unfold. Increasing the knowledge of basic developmental and signaling principles that are controlled by endocannabinoids will provide important insights into the molecular mechanisms that establish functional neuronal circuits in the brain. Moreover, determining the molecular basis of permanent modifications to cellular structure and intercellular communication imposed by cannabis smoking during pregnancy will provide novel therapeutic targets for alleviating pathogenic changes in affected offspring. Here, we summarize recent findings regarding the ontogeny of the endocannabinoid system in neurons that sculpt the temporal and spatial diversity of cellular functions during CNS development.

Animals↗

In vitro cytotoxicity of the organophosphorus pesticide parathion to FG-9307 cells.

FG-9307, a cell line derived from a gill of the flounder, Paralichthys olivaceus, was used to determine the cytotoxic effects of the organophosphorus (OP) pesticide parathion. Cytotoxicity was measured by three endpoint systems: neutral red (NR) uptake assay, tetrazolium (MTT) assay and cell protein assay. The lowest concentration of parathion tested (1 microg/ml) was toxic and there was no significant difference in cytotoxic effects among the three assays. The FG-9307 cell line is a suitable bioindicator for the screening of the acute toxicities of parathion. The fine structures of the cells were also studied. Ultrastructures were markedly altered by parathion, as evidenced by dilation of nuclear membranes and mitochondrial cristae and by the presence of lysosomes with engulfed particles. With the increase of the parathion concentration, the damage degree of the cellular structures was more serious. At the highest concentration tested (15 microg/ml), there were few visible organelles, although such changes in cell morphology were not observed under a light microscope. Apparently, this is the unnoted report of marine fish cell line used for the evaluation of the acute in vitro cytotoxicity of parathion.

Animals↗

Changes in carotenoids during processing and storage of foods.

Being highly unsaturated, carotenoids are susceptible to isomerization and oxidation during processing and storage of foods. Isomerization of trans-carotenoids to cis-carotenoids, promoted by contact with acids, heat treatment and exposure to light, diminishes the color and the vitamin A activity of carotenoids. The major cause of carotenoid loss, however, is enzymatic and non-enzymatic oxidation, which depends on the availability of oxygen and the carotenoid structure. It is stimulated by light, heat, some metals, enzymes and peroxides and is inhibited by antioxidants. Data on percentage losses of carotenoids during food processing and storage are somewhat conflicting, but carotenoid degradation is known to increase with the destruction of the food cellular structure, increase of surface area or porosity, length and severity of the processing conditions, storage time and temperature, transmission of light and permeability to O2 of the packaging. Contrary to lipid oxidation, for which the mechanism is well established, the oxidation of carotenoids is not well understood. It involves initially epoxidation, formation of apocarotenoids and hydroxylation. Subsequent fragmentations presumably result in a series of compounds of low molecular masses. Completely losing its color and biological activities, the carotenoids give rise to volatile compounds which contribute to the aroma/flavor, desirable in tea and wine and undesirable in dehydrated carrot. Processing can also influence the bioavailability of carotenoids, a topic that is currently of great interest.

Carotenoids↗

The temporal architecture of eukaryotic growth.

Coherence of the time structure of growing organisms depends on a metronome-like orchestration. In a continuously perfused culture of Saccharomyces cerevisiae the redox state of the cell shows a temperature-compensated oscillation manifest in respiratory cycles, which are measured by continuous and non-invasive electrodes of probes such as dissolved oxygen and probes such as fluorometric NAD(P)H. Although the entire transcriptome exhibits low-amplitude oscillatory behaviour, transcripts involved in the vast majority of metabolism, stress response, cellular structure, protein turnover, mRNA turnover, and DNA synthesis are amongst the top oscillators and their orchestration occurs by an intricate network of transcriptional regulators. Therefore cellular auto-dynamism is a function of a large ensemble of excitable intracellular components of that self-organized temporally and spatially that encompasses mitochondrial, nuclear, transcriptional and metabolic dynamics, coupled by cellular redox state.

Biological Evolution↗

Mechanical tuning characteristics of the hearing organ measured at the sensory cells in the gerbil temporal bone preparation.

The micromechanical behaviour of the inner ear in response to sound stimulation was investigated in an in vitro preparation of the gerbil temporal bone. Using laser heterodyne interferometry it was possible to measure the vibration responses directly at the level of the sensory and supporting cells within the hearing organ rather than from the underlying basilar membrane as has been done in previous studies. There was a tuned mechanical response of the cellular structures within the hearing organ. The resonance frequency measured at cells in the apical (third) turn was around 200Hz. The frequency of the mechanical tuning varied along the length of the cochlea. In the second turn the resonance frequency was around 500-700Hz. The cellular response in the second turn was more sharply tuned as compared to the response in the apical turn. In both cochlear turns the amplitude of the vibratory response changed with the cellular location radially across the hearing organ.

Acoustic Stimulation↗

Discovery of the 'porosome'; the universal secretory machinery in cells.

The release of neurotransmitters at the nerve terminal for neurotransmission, release of insulin from beta-cells of the endocrine pancreas for regulating blood glucose levels, the release of growth hormone from GH cells of the pituitary gland to regulate body growth, or the expulsion of zymogen from exocrine pancreas to digest food, are only a few examples of key physiological processes made possible by cell secretion. It comes as no surprise that defects in cell secretion are the cause for numerous diseases, and have been under intense investigation for over half century. Only in the last decade, the molecular machinery and mechanism of cell secretion has become clear. Cell secretion involves the docking and transient fusion of membrane-bound secretory vesicles at the base of plasma membrane structures called porosomes, and the regulated expulsion of intravesicular contents to the outside, by vesicle swelling. The discovery of the porosome in live cells, its morphology and dynamics at nanometer resolution and in real time, its isolation, its composition, and its structural and functional reconstitution in lipid membrane, are complete. The molecular mechanism of secretory vesicle fusion at the base of porosomes, and the regulated expulsion of intravesicular contents during cell secretion, are also resolved. In this minireview, the monumental discovery of the porosome, a new cellular structure at the cell plasma membrane, is briefly discussed.

Amino Acid Sequence↗

Noninvasive imaging of oral premalignancy and malignancy.

Early detection of cancer and its precursors remains the best way to ensure patient survival and quality of life. Our specific aim is to test a multimodality approach to noninvasive diagnostics of oral premalignancy and malignancy. In the hamster cheek pouch model (120 hamsters), in vivo optical coherence tomography (OCT) and optical Doppler tomography (ODT) map epithelial, subepithelial, and vascular change throughout carcinogenesis. In vivo multiwavelength multiphoton (MPM) and second-harmonic generated (SHG) fluorescence techniques provided parallel data on surface and subsurface tissue structure, specifically collagen presence and structure, cellular presence, and vasculature. Images are diagnosed by two blinded, prestandardized investigators using a scale from 0 to 6 for all modalities. After sacrifice, histopathology is evaluated on a scale of 0 to 6. Imaging data are reproducibly obtained with good accuracy. Carcinogenesis-related structural and vascular changes are clearly visible to tissue depths of 2 mm. Sensitivity (OCT/ODT alone, 71 to 88%; OCT+MPMSHG, 79 to 91%) and specificity (OCT alone, 62 to 83%; OCT+MPMSHG, 67 to 90%) compare well with conventional techniques. Our conclusions are that OCT/ODT and MPM/SHG are promising noninvasive in vivo diagnostic modalities for oral dysplasia and malignancy.

Animals↗

2,2'-thiodiethanol: a new water soluble mounting medium for high resolution optical microscopy.

The use of high numerical aperture immersion lenses in optical microscopy is compromised by spherical aberrations induced by the refractive index mismatch between the immersion system and the embedding medium of the sample. Especially when imaging >10 micro m deep into the specimen, the refractive index mismatch results in a noticeable loss of image brightness and resolution. A solution to this problem is to adapt the index of the embedding medium to that of the immersion system. Unfortunately, not many mounting media are known that are both index tunable as well as compatible with fluorescence imaging. Here we introduce a nontoxic embedding medium, 2,2'-thiodiethanol (TDE), which, by being miscible with water at any ratio, allows fine adjustment of the average refractive index of the sample ranging from that of water (1.33) to that of immersion oil (1.52). TDE thus enables high resolution imaging deep inside fixed specimens with objective lenses of the highest available aperture angles and has the potential to render glycerol embedding redundant. The refractive index changes due to larger cellular structures, such as nuclei, are largely compensated. Additionally, as an antioxidant, TDE preserves the fluorescence quantum yield of most of the fluorophores. We present the optical and chemical properties of this new medium as well as its application to a variety of differently stained cells and cellular substructures.

Animals↗

[Three-dimensional reconstruction of cytoskeletons in rat salivary glands using confocal fluorescence microscopy and volume-rendering computer graphics].

A new reconstruction technique for the demonstration of three-dimensional architectural details from biological fluorescent specimens was used to determine the precise spacial organization of F-actin in rat sublingual glands. F-actin was stained with NBD-phallacidin in thick (20-30 microns) frozen sections and observed with a confocal laser scanning microscope to obtain thin (-1 micron) optically sectioned images. A series of optical sections taken successively at different focal levels in steps of 1 micron was then reconstructed with volume-rendering computer graphics. The rendered images viewed from several angles (e.g. top, bottom, sides) clearly revealed the presence of F-actin fluorescence along the hexagonal framework of the junctional complex within the acini and in the cytoplasm of star-shaped myoepithelial cells encircling the acini. The detailed structures and the topographical relations between the junctional complex and myoepithelial cells were more impressively observed when the rendered images were displayed by motion picture. These images are free from any artifacts caused by the mechanical sectioning and revealed well the delicate and complicated profiles of cellular structures that previously have been almost impossible to demonstrate by conventional reconstruction techniques. We expect that the combination of confocal microscopy and volume rendering will permit the dynamic observation of previously unseen phenomena in three dimensions, such as the behavior of biologically active molecules in living cells.

Animals↗

Cell specialization in the small intestinal epithelium of adult Xenopus laevis: structural aspects.

The structure, cellular composition and organization of the small intestinal epithelium of adult Xenopus laevis are described. The mucosa is thrown into a system of longitudinal folds and is composed mainly of columnar cells and goblet cells. Leucocytes comprise about 28% of the mucosal cells. The degree of structural specialization of columnar cells varies according to their position in the folds. Cells at the crest of the folds display the greastest degree of specialization, evidenced by the degree of development of their inter- and intracellular membrane systems. Cells in the trough regions between the folds are much less specialized. These cell types are structurally analogous to mammalian villous and crypt cells respectively, although they are not segregated to the same degree.

Animals↗

Taking X-ray diffraction to the limit: macromolecular structures from femtosecond X-ray pulses and diffraction microscopy of cells with synchrotron radiation.

Recent work is extending the methodology of X-ray crystallography to the structure determination of noncrystalline specimens. The phase problem is solved using the oversampling method, which takes advantage of "continuous" diffraction patterns from noncrystalline specimens. Here we review the principle of this newly developed technique and discuss the ongoing experiments of imaging nonperiodic objects, such as cells and cellular structures, using coherent and bright X rays produced by third-generation synchrotron sources. In the longer run, the technique may be applicable to image single biomolecules using anticipated X-ray free electron lasers. Here, computer simulations have so far demonstrated two important steps: (a) by using an extremely intense femtosecond X-ray pulse, a diffraction pattern can be recorded from a macromolecule before radiation damage manifests itself; and (b) the phase information can be retrieved in an ab initio fashion from a set of calculated noisy diffraction patterns of single protein molecules.

Algorithms↗

Exploring RPE as a source of photoreceptors: differentiation and integration of transdifferentiating cells grafted into embryonic chick eyes.

PURPOSE: To study the possibility of generating photoreceptors through programming RPE transdifferentiation by examining cell differentiation after transplantation into the developing chick eye. METHODS: RPE was isolated, and the cells were dissociated, cultured, and guided to transdifferentiate by infection with retrovirus expressing neuroD (RCAS-neuroD), using RCAS-green fluorescence protein (GFP) as a control. The cells were then harvested and microinjected into the developing eyes of day 5 to day 7 chick embryos, and their development and integration were analyzed. RESULTS: Cells from the control culture integrated into the host RPE. When grafted cells were present in large number, multilayered RPE-like tissues were formed, and the extra tissues consisted of grafted cells and host cells. None of the cells from the control culture expressed photoreceptor-specific genes. In contrast, most cells from RCAS-neuroD-infected culture remained depigmented. A large number of them expressed photoreceptor-specific genes, such as visinin and opsins. Antibodies against red opsin decorated the apical tips and the cell bodies of the grafted, transdifferentiating cells. In the subretinal space, visinin(+) cells aligned along the RPE or an RPE-like structure. When integrated into the host outer nuclear layer, grafted cells emanated elaborate, axonal arborization into the outer plexiform layer of the host retina. CONCLUSIONS: Cultured RPE cells retained their remarkable regenerative capabilities. Cells guided to transdifferentiate along the photoreceptor pathway by neuroD developed a highly ordered cellular structure and could integrate into the outer nuclear layer. These data suggest that, through genetic programming, RPE cells could be a potential source of photoreceptor cells.

Animals↗

Sperm autoantigens and fertilization III. Ultrastructural localization of guinea pig autoantigens.

Guinea pig sperm autoantigens have been localized by direct and indirect immunoferritin techniques in (1) plasma membrane over the entire sperm head, (2) acrosomal contents, (3) fibrous sheath and outer dense fibers of the tail filament, and (4) the inner acrosomal membrane of 50% of acrosome reacted spermatozoa. These cellular structures are known to be involved in guinea pig sperm rouleaux formation, acrosome reaction, interaction of acrosome-reacted sperm with zona pellucida and with the vitellus of guinea pig ova. Since IgG and Fab of autoantiserum to guinea pig spermatozoa have been shown to interfere with these cellular reactions, this study provides further evidence, albeit indirect, that sperm autoantigens are involved in these cellular events.

Acrosome↗

Histone phosphorylation during sea urchin development.

Studies on histone phosphorylation during transitions in chromatin structure occurring in vivo during spermatogenesis and early embryogenesis in sea urchins are reviewed and evaluated in the light of recent studies on histone phosphorylation occurring during chromatin synthesis in frog egg extracts in vitro and evidence that protein kinases and phosphatases play direct roles in the regulation of cellular structure. Sperm-specific histone variants Sp H1 and Sp H2B are maintained as phosphorylated derivatives N and O/P throughout spermatogenesis and early embryogenesis and egg specific histone variants CS H1 and CS H2A are phosphorylated during early embryogenesis. These developmental correlations provide clues about the roles of histone phosphorylation in control of chromatin structure in vivo and provide a basis for the interpretation of data obtained from in-vitro sperm chromatin remodeling in egg extracts and from biochemical studies on the effects of histone phosphorylation on DNA binding. The potential consequences for chromatin structure of the various histone phosphorylation events observed in sea urchins and frog egg extracts are discussed.

Animals↗

A protein kinase bound to the projection portion of MAP 2 (microtubule-associated protein 2).

In previous work we have demonstrated that the microtubule-associated protein 2 (MAP 2) molecule consists of two structural parts. One part of the molecule, referred to as the assembly-promoting domain, binds to the microtubule surface and is responsible for promoting microtubule assembly; the other represents a filamentous projection observed on the microtubule surface that may be involved in the interaction of microtubules with other cellular structures. MAP 2 is known to be specifically phosphorylated as the result of a protein kinase activity that is present in microtubule preparations. We have now found that the activity copurifies with the projection portion of MAP 2 itself. Kinase activity coeluted with MAP 2 when microtubule protein was subjected to either gel- filtration chromatography on bio-gel A-15m or ion-exchange chromatography on DEAE- Sephadex. The activity was released from microtubules by mild digestion with chymotrypsin in parallel with the removal by the protease of the MAP 2 projections from the microtubule surface. The association of the activity with the projection was demonstrated directly by gel filtration chromatography of the projections on bio-gel A-15m. Three protein species (M(r) = 39,000, 55,000, and 70,000) cofractionated with MAP 2, and two of these (M(r) = 39,000 and 55,000) may represent the subunits of an associated cyclic AMP- dependent protein kinase. The projection-associated activity was stimulated 10-fold by cyclic AMP and was inhibited more than 95 percent by the cyclic AMP-dependent protein kinase inhibitor from rabbit skeletal muscle. It appeared to represent the only significant activity associated with microtubules, almost no activity being found with tubulin, other MAPs, or the assembly-promoting domain of MAP 2, and was estimated to account for 7-22 percent of the total brain cytosolic protein kinase activity. The location of the kinase on the projection is consistent with a role in regulating the function of the projection, though other roles for the enzyme are also possible.

Animals↗

Comparison of the antibiotic resistance mechanisms in a gram-positive and a gram-negative bacterium by gene networks analysis.

Nowadays, the emergence of some microbial species resistant to antibiotics, both gram-positive and gram-negative bacteria, is due to changes in molecular activities, biological processes and their cellular structure in order to survive. The aim of the gene network analysis for the drug-resistant Enterococcus faecium as gram-positive and Salmonella Typhimurium as gram-negative bacteria was to gain insights into the important interactions between hub genes involved in key molecular pathways associated with cellular adaptations and the comparison of survival mechanisms of these two bacteria exposed to ciprofloxacin. To identify the gene clusters and hub genes, the gene networks in drug-resistant E. faecium and S. Typhimurium were analyzed using Cytoscape. Subsequently, the putative regulatory elements were found by examining the promoter regions of the hub genes and their gene ontology (GO) was determined. In addition, the interaction between milRNAs and up-regulated genes was predicted. RcsC and D920_01853 have been identified as the most important of the hub genes in S. Typhimurium and E. faecium, respectively. The enrichment analysis of hub genes revealed the importance of efflux pumps, and different enzymatic and binding activities in both bacteria. However, E. faecium specifically increases phospholipid biosynthesis and isopentenyl diphosphate biosynthesis, whereas S. Typhimurium focuses on phosphorelay signal transduction, transcriptional regulation, and protein autophosphorylation. The similarities in the GO findings of the promoters suggest common pathways for survival and basic physiological functions of both bacteria, including peptidoglycan production, glucose transport and cellular homeostasis. The genes with the most interactions with milRNAs include dpiB, rcsC and kdpD in S. Typhimurium and EFAU004_01228, EFAU004_02016 and EFAU004_00870 in E. faecium, respectively. The results showed that gram-positive and gram-negative bacteria have different mechanisms to survive under antibiotic stress. By deciphering their intricate adaptations, we can develop more effective therapeutic approaches and combat the challenges posed by multidrug-resistant bacteria.

Anti-Bacterial Agents↗